Amino acid and hydroxyamino acid transporter compounds for therapeutic applications, process and use

ABSTRACT

Amino acid and hydroxyamino acid transporter compounds are provided, in which an amino acid or hydroxyamino acid as a carrier is linked via an ester linkage to a therapeutic compound, and having one of the general formulae: 
     
         NH.sub.3.sup.+ --AA--COO--Z.sub.1                          I 
    
     
         NH.sub.3.sup.+ --HAA--OOC--Z.sub.2                         II 
    
     in which AA represents the amino acid or hydroxyamino acid, HAA represents the hydroxyamino acid, and Z 1  and Z 2  represent a therapeutic compound, or a linking compound attached to COOH or OH of the hydroxyamino acid and to the therapeutic compound, as well as a process for preparing the same, and a process for administration of the same to animals, to obtain the benefit of the therapeutic effect of the therapeutic compound.

Disorders in animals including man are treated by administration of atherapeutic agent, whose therapeutic effect is obtained only uponabsorption and utilization in the metabolic system of the animal. If thetherapeutic agent cannot be absorbed readily in the metabolic system,its capabilities and effectiveness naturally are greatly diminished.

A prerequisite for absorbability of a therapeutic agent is that itpenetrate biological barriers, such as the blood brain barrier or cellmembranes. The ability of a therapeutic agent to penetrate such barriersis greatly influenced by the chemical and physical properties of theagent, and also upon the integrity and chemical nature of the barrier ormembrane.

In general, hydrophilic therapeutic agents are readily absorbed andlipophilic therapeutic agents are poorly absorbed by the animal body.Lipophilic therapeutic agents penetrate biological membranes at slowrates of transport, generally by passive mechanisms, in part becauseactive transport mechanisms across biological membranes and similarbarriers exist only for hydrophilic compounds. Therapeutic compoundshaving a high molecular weight and/or a complex spatial arrangement ofthe molecule, often features of pharmacologically active compoundsbecause spatial structure is frequently most important in therapeuticactivity, also are transported only slowly across biological membranes.

Traditionally, research efforts to improve transport have endeavoured tomanipulate the molecule of the therapeutic agent by adding groups thatenhance solubility and therefore bioavailability of the compound. Suchan approach has several inherent disadvantages. First, a new compound iscreated, of uncertain therapeutic effectiveness. Since the compound isnew, existing regulatory requirements require a complete and costlyreevaluation of the new compound's effectiveness and toxicity, includingcarcinogenic potential, and this is of course involves considerableexpense and effort, as well as time, and may be unsuccessful. Completionof the reevaluation may only show that the compound has a substantiallydifferent therapeutic effectiveness in animals, because of the alteredstructure/activity relationship. In many instances, toxicity problemsarise. Such new compounds may be carcinogenic, even though thepredecessor molecule is not.

In accordance with the present invention, a new approach is madeutilizing an amino acid or hydroxyamino acid as a transporter for knowntherapeutic agents, linking the amino acid or hydroxyamino acid to thetherapeutic agent by way of an ester linkage between the amino acid orhydroxyamino acid molecule and the molecule of the therapeutic agent. Apreferred amino acid is lysine, having two amino groups with a positivecharge and a considerable hydrophilic character, and a preferredhydroxyamino acid is serine. The amino acid or hydroxyamino acid acts asa carrier for the therapeutic compound, in transport of the compoundacross biological barriers, including cell membranes and the blood brainbarrier. After passage of the barrier, the new compound enters themetabolic system of the animal, where the ester linkage is destroyed bynaturally-occurring esterases present in the animal, thus regeneratingfree amino acid or hydroxyamino acid and the known therapeutic compound.Since the known therapeutic compound is of known therapeuticeffectiveness, no problems with existing regulatory requirements arise.Similarly, since amino acids and hydroxyamino acids as a class,including in particular lysine and serine, are known to be safe andnontoxic, no problems arise because of its presence. The result is aconsiderably enhanced capability of the therapeutic agent to be absorbedin the metabolic system of the animal, without any complicating sideeffects.

The amino acid and hydroxyamino acid transporter compounds in accordancewith the invention can link to the therapeutic compound via a carboxylicacid group COOH of the amino acid or hydroxyamino acid, or via ahydroxyl group of the hydroxyamino acid, and accordingly have one of thefollowing general formulae:

    NH.sub.3.sup.+ --AA--COO--Z                                I

(linked via a COOH group of the amino acid or hydroxyamino acid)

    NH.sub.3.sup.+ --HAA--OOC--Z                               II

(linked via a hydroxyl group of the hydroxyamino acid)

In the above formulae, AA represents the amino acid or hydroxyaminoacid, HAA represents the hydroxyamino acid, radical linked to COO-- orO--; Z represents (1) therapeutic compound, or (2) a therapeuticcompound and a linking compound attached to the COOH or OH of the aminoacid or hydroxyamino acid, and via another ester group to thetherapeutic compound.

Any therapeutic compound having a hydroxyl, carboxylic acid or acylgroup can form an ester linkage with (a) the amino acid or hydroxyaminoacid directly, by way of a carboxy group of the amino or hydroxyaminoacid, or (b) a linking compound having a carboxyl group, in the case ofa hydroxyl group on the therapeutic compound, or with (c) a hydroxylgroup on the hydroxyamino acid, or (d) a hydroxyl group on the linkingcompound, with a carboxylic or acyl group on the therapeutic compound.

The linking compound provides a way of forming the ester linkage withthe amino or hydroxyamino acid of a therapeutic compound with acarboxylic acid or acyl group, since the linking compound can have twohydroxyl groups, or one hydroxyl and one acid group, which can formeaster linkages, one with carobxylic acid group of the amino orhydroxyamino acid, and one with the carboxylic acid or acyl or hydroxylgroup of the therapeutic compound. These compounds constitute a subclasswithin Formula I that have one of the general formulae:

    NH.sub.3.sup.+ --AA--COO--Y.sub.1 --OOC--Z.sub.2           III

    NH.sub.3.sup.+ --AA--COO--Y.sub.2 --COO--Z.sub.1           IV

The hydroxyl group of a hydroxyamino acid can also be linked to acarboxylic acid group of a linking compound, which has anothercarboxylic acid group or one hydroxyl group that can be linked to thehydroxyl or carboxylic acid or acyl group of a therapeutic compound.These compounds constitute a further subclass within formula II thathave one of the general formulae:

    NH.sub.3.sup.+ --HAA--OOC--Y.sub.3 --COO--Z.sub.1          V

    NH.sub.3.sup.+ --HAA--OOC--Y.sub.2 --OOC--Z.sub.2          VI

In the above formulae AA represents an amino acid, HAA represents ahydroxyamino acid, Z₁ represents a therapeutic compound having a hydroxygroup; and Z₂ represents a therapeutic compound having a carboxylic acidor acyl group, Y₁ a linking compound having two hydroxyl groups, Y₂ alinking compound having one hydroxyl and one carboxylic acid or acylgroup, and Y₃ a linking compound having two carboxylic acid groups. Anexample of Y₁ is two tyrosine groups linked by a peptide linkage, as inthe following example of lysine-tyrosine-linked-tryptophane compounds ofFormula VII: ##STR1##

In this example, AA is lysine, Y is a double tyrosine molecule linked bya peptide linkage CONH and Z is tryptophane, linked to Y by way of thecarboxylic acid group.

The transporter compounds of the invention can utilize any amino acid,as a class, of which hydroxyamino acids are a preferred subclass. Theamino acids have at least one amino group, and at least one carboxylicacid group, and the hydroxyamino acids have at least one amino group, atleast one carboxylic acid group and one hydroxyl group. The number ofcarbon atoms is not critical, except that larger molecules can displaydiminished solubility in pharmaceutically acceptable solvents; usuallythe amino acid has from two to about twenty carbon atoms.

A plurality of amino groups and a plurality of carboxylic acid groupsand a plurality of hydroxyl groups are no detriment, since all threetypes of groups are hydrophilic, and thus increase hydrophilicity. Theamino acids can have up to five amino groups, up to five carboxylic acidgroups, and up to five hydroxyl groups.

The relative positions of the amino, COOH and OH (if any) groups on themolecule are not critical, either, but it is usually preferred that theybe α, β, γ or δ to each other, since proximity may be advantageous.

The amino acids and hydroxyamino acids can be aliphatic, cycloaliphatic,mixed aliphatic-aromatic, mixed aliphatic-cycloaliphatic, or mixedcycloaliphatic-aromatic.

Exemplary aliphatic amino acids include: glycine, glycyl glycine, α- andβ-alanine, Lysine, tryptophane, valine, norvaline, leucine, isoleucine,norleucine, cystine, methionine, arginine, asparagine, creatine,glutamine, arginine, α-aminobutyric acid, α,ε-diaminopimelic acid,lanthionine, djenkolic acid, γ-methyleneglutamic acid, α,γ-diaminobutyric acid, ornithine, citrulline, and canavarine.

Exemplary cycloaliphatic amino acids include: proline, histidine.

Exemplary mixed aliphatic-aromatic amino acids include: thyroxine,diiodothyroxine and phenylalanine.

Exemplary mixed aliphatic cycloaliphatic amino acids include: histidine.

Exemplary mixed aromatic-cycloaliphatic amino acids include:tryptophane.

Exemplary aliphatic hydroxyamino acids include: hydroxyamino butyricacid, δ-hydroxy lysine, hyroxyvaline tyrosine, serine, threonine,cysteine (with SH instead of OH), aspartic acid, hydroxyglutamic acid,allothreonine, α- and β-thiolvaline, diaminetrihydroxy dodecanoic acidand diaminodihydroxy suberic acid.

Exemplary cycloaliphatic hydroxyamino acids include: hydroxyproline.

Exemplary mixed aliphatic-aromatic hydroxyamino acids include: tyrisine,thyroxine, diiodo tyrosine, and surinamine.

The amino acid and hydroxyamino acid transporter compounds of thepresent invention are readily prepared by conventional esterificationprocedures, which are well known in the art. The following Example showsthe preparation of O-L-lysyl-L-tyrosine.

Example I

The reaction sequence is shown below:

    ______________________________________                                         ##STR2##                                                                      ##STR3##                                                                      ##STR4##                                                                      ##STR5##                                                                      ##STR6##                                                                     BOC.sub.2LysOnSu                                                                           Nα-Nε-bis-t-BOCL-lysine                                         Nhydroxysuccinimide ester                                        BOCTyrOBZl   Nα-t-BOCL-tyrosine benzyl ester                            NaH          sodium hydride                                                   DMF          N,Ndimethylformamide                                             EtOAC        ethyl acetate                                                    ______________________________________                                    

N-α-t-BOC-O-(Nα'-Nε'-bis-t-BOC-L-lysyl)-L-tyrosine benzyl ester³

Into a 250 ml 3-neck, round-bottom flask equipped with magnetic stirrer,thermometer, drying tube and cooling bath, were charged 10 g (27.0mmole) BOC-Tyr-OBZl (1) and 125 ml DMF. The solution was cooled to 0°C., and 1.1 g (27.5 mmole) NaH (60% oil dispersion) was added in oneportion. The solution/suspension was stirred one hour at -10°-0° C.,then 12.0 g (27.1 mmole) BOC₂ LysONSu (2) was added and the reactionstirred overnight at room temperature. The mixture was poured into 250ml ice/H₂ O, then extracted with CH₂ Cl₂ (3×350 ml). The combinedorganic extracts were washed with 1N NaOH (2×500 ml), H₂) (3×500 ml),sat. NaCl (1×500 ml) and dried over Na₂ SO₄. The organic phase was thenevaporated in vacuo to yield 16.3 g (86%) of a white solid, pure by TLCCHCl₃ :CH₂ OH:CH₂ CO₂ H-9: 1:1. IR(cm⁻¹):3380, 3390(NH); 1700, 1710,1745, 1768 (c=0). NMR(ppm):2.43 (s, 9H), 2.47 (s, 9H), 2.50 (s, 9H0,7.03 (M, 4H), 7.34 (M, 5H).

N-α-t-BOC-O-(Nα'-Nε'-bis-t-BOC-L-lysyl0-L-tyrosine ⁴

Into a 250 ml 3-neck, round-bottom flask equipped with a magneticstirrer and gas inlet and outlet, were charged 16.1. g³, 90 ml DMF, 4.50g HCO₂ NH₄ and 4.5 g 10% Pd/c. After scrubbing with N₂, H₂ was bubbledthrough the mixture at atmospheric pressure. After 5 minutes, thereduction was complete and the catalyst was filtered off, and thefiltrate was partitioned between CH₂ Cl₂ /1NHCl (500 ml/500 ml). Theorganic phase was washed with 1NHCl (1×400 ml), H₂ O (4×500 ml).saturated NaCl (1×500 ml) and dried over Na₂ SO₄. After evaporation ofthe CH₂ Cl₂ and trituration with hexane, 13.8 g (97%) of ⁴ was obtained.TLC: EtOAc:CH₃ CO₂ H:hexane-20: 10:1 showed one spot. IR (cm⁻¹):3385(NH); 1675, 1700, 1728, 1770(c=0).

O-L-Lysyl-L-tyrosine·3 HCL⁵

A solution of 13.5 g (22.2 mmole)⁴ in liter EtOAc was placed in a 2liter, 3 neck, round-bottom flask equipped with magnetic stirrer, gasinlet tube, and drying tube. Anhydrous HCl was bubbled through thesolution for 30 minutes. A precipitate appeared after about 15 minutes.The precipitate was filtered off, washed with EtOAc and dried in vacuoto yield 9.3 g (100%) of hygroscopic product. TLC--nBuOH: CH₂ CO₂ H: H₂O-4: 1: 1 Rf=0.2, trace 0.8. IR(cm⁻¹): 1950 (broad NH₃ ⁺); 1750, 1700(c=0).

The invention is broadly applicable to any therapeutic substances havingone or more free hydroxyl groups capable of reacting with the carboxylicacid group or groups of the amino or hydroxyamino acid, as in Formula Iabove, or with one or more free carboxylic acid or acyl groups capableof reacting with the hydroxyl group of α-hydroxyamino acid, as inFormula II above, or of an intermediary linking compound having at leasttwo hydroxyl or carboxylic acid groups, of which one is linked to theamino acid or hydroxyamino acid as in Formulae III, IV, and V above.

The following is a brief list of classes of known therapeutic agentswhich can be so linked, and whose absorptivity in the metabolic systemof animals is thereby greatly facilitated:

(1) amino acids

(2) depsipeptides

(3) peptides

(4) polypeptides

(5) proteins

(6) psychotropic medications known as

(1) tranquilizers

(2) sedatives

(3) antidepressants

(4) neuroleptics

(5) hypnotics

(6) muscle relaxants

(7) anticonvulsants

(8) analgesics

(9) analeptics

(10) anesthetics

(11) antiParkinsonian agents

(12) CNS stimulants

(13) psychostimulants

(7) antiasthma compounds

(8) antispasmotics

(9) anorexics

(10) cardiovascular agents

(1) antiarthymics

(2) antihypertensives

(3) cardiac glycosides

(4) antidiuretics

(5) antimigraines

(11) antibacterials and antiseptics

(12) antibiotics

(13) antineoplastic drugs

(14) anticoagulants

(15) antidiabetic agents

(16) antidiarrheals

(17) antidotes

(18) antifungal agents

(19) antihistamines

(20) antiherpes (and other antiviral)

(21) anti-inflammatory agents

(22) antimetabolites

(23) antimalarials

(24) antiemetics

(25) antiparasitics

(26) antipruiritics

(27) antipyretics

(28) antispasmotics, anticholinergics

(29) biologicals

(30) bronchodilators

(31) calcium preparations

(32) antihyperlipidemics

(33) contraceptives

(34) cough and cold preparations

(35) decongestants

(36) dental preparations

(37) dermatologicals

(38) diagnostics

(39) dietary supplements

(40) hormones

(41) immunosuppressives

(42) ophthalmologicals

(43) parasympatholytics

(44) parasmypathomimetics

(45) prostaglandins

The following lysine transporter compounds are exemplary, in which (1)therapeutic compounds are indicated by the symbol "X", where "X" iseither

1. vincristine

2. vinblastine

3. methotrexate

4. daunorubicin

5. bleomycin

6. cytosine arabinoside

7. thiotepa

8. dactinomycin

9. doxorubicin

10. mithramycin

11. mitomycin

12. azauridine

13. idoxuridine

14. deoxyuridine

15. paracetemol

16. calicylic acid

17. acetylsalicylic acid

18. lecithin

19. lecithen-peptide-tryptophan

20. cromoglycate

21. doxepin

22. amoxapine

23. thiothixene

24. lorazepam

25. oxazepam

26. temazepam

27. normetazepam

28. leucovorin

29. tyrosine

30. tryptophan

31. dopamine

32. 5-hydroxytryptophan

33. 5-hydroxytryptamine

34. serotonin

and in which (2) carrier compounds are indicated by the symbol "R",where "R" is either

1. lysine

2. serine

3. seryllysine

4. lysyllysine

5. lysylserine

6. lysylseryllysine

7. 2,6 diaminohexanol

8. delta hydroxylysine

9. lactic acid

10. 2,4 diamino butanol

EXEMPLARY COMPOUNDS

1. L-R-ester-L-X

2. D-R-ester-L-X

3. D-R-ester-D-X

4. L-R-ester-D-X

5. L-try-ester-L-tyr-peptide-L-tyr-ester-L-R

6. D-try-ester-L-tyr-peptide-L-tyr-ester-L-R

7. L-try-ester-D-tyr-peptide-L-tyr-ester-L-R

8. L-try-ester-L-tyr-peptide-D-tyr-ester-L-R

9. L-try-ester-L-tyr-peptide-L-tyr-ester-D-R

10. D-try-ester-D-tyr-peptide-L-tyr-ester-L-R

11. D-try-ester-L-tyr-peptide-D-tyr-ester-L-R

12. D-try-ester-L-tyr-peptide-L-tyr-ester-D-R

13. L-try-ester-D-tyr-peptide-D-tyr-ester-L-R

14. L-try-ester-D-tyr-peptide-L-tyr-ester-D-R

15. L-try-ester-L-tyr-peptide-D-tyr-ester-D-R

16. D-try-ester-D-tyr-peptide-D-tyr-ester-L-R

17. D-try-ester-D-tyr-peptide-L-tyr-ester-D-R

18. D-try-ester-L-tyr-peptide-D-tyr-ester-D-R

19. L-try-ester-D-tyr-peptide-D-tyr-ester-D-R

20. D-try-ester-D-tyr-peptide-D0tyr-ester-D-R

21. L-R-ester-L-tyr-peptide-L-tyr-ester-L-try

22. D-R-ester-L-tyr-peptide-L-tyr-ester-L-try

23. L-R-ester-D-tyr-peptide-L-tyr-ester-L-try

24. L-R-ester-L-tyr-peptide-D-tyr-ester-L-try

25. L-R-ester-L-tyr-peptide-L-tyr-ester-D-try

26. D-R-ester-D-tyr-peptide-L-tyr-ester-L-try

27. D-R-ester-L-tyr-peptide-D-tyr-ester-L-try

28. D-R-ester-L-tyr-peptide-L-tyr-ester-D-try

29. L-R-ester-D-tyr-peptide-D-tyr-ester-L-try

30. L-R-ester-D-try-peptide-L-tyr-ester-D-try

31. L-R-ester-L-tyr-peptide-D-tyr-ester-D-try

32. D-R-ester-D-tyr-peptide-D-tyr-ester-L-try

33. D-R-ester-D-tyr-peptide-L-tyr-ester-D-try

34. D-R-ester-L-tyr-peptide-D-tyr-ester-D-try

35. L-R-ester-D-tyr-peptide-D-tyr-ester-D-try

36. D-R-ester-D-tyr-peptide-D-tyr-ester-D-try

37. L-try-ester-L-tyr-ester-L-R

38. D-try-ester-L-tyr-ester-L-R

39. L-try-ester-D-tyr-ester-L-R

40. L-try-ester-L-tyr-ester-D-R

41. D-try-ester-D-tyr-ester-L-R

42. D-try-eseter-L-tyr-ester-D-R

43. L-try-ester-D-tyr-ester-D-R

44. D-try-ester-F-tyr-ester-D-R

45. L-R-ester-L-tyr-ester-L-try

46. D-R-ester-L-tyr-ester-L-try

47. L-R-ester-D-tyr-ester-L-try

48. L-R-ester-L-tyr-ester-D-try

49. D-R-ester-D-tyr-ester-L-try

50. D-R-ester-L-tyr-ester-D-try

51. L-R-ester-D-tyr-ester-D-try

52. D-R-ester-D-tyr-ester-D-try

53. L-tyr-ester-L-try-peptide-L-try-ester-L-R

54. D-try-ester-L-try-peptide-L-try-ester-L-R

55. L-tyr-ester-D-try-peptide-L-try-ester-L-R

56. L-tyr-ester-L-try-peptide-D-try-ester-L-R

57. L-tyr-ester-L-try-peptide-L-try-ester-D-R

58. D-tyr-ester-d-try-peptide-L-try-ester-L-R

59. D-tyr-ester-L-try-peptide-D-try-ester-L-R

60. D-tyr-ester-L-try-peptide-L-try-ester-D-R

61. L-tyr-ester-D-try-peptide-D-try-ester-L-R

62. L-tyr-ester-D-try-peptide-L-try-ester-D-R

63. L-tyr-ester-L-try-peptide-D-try-ester-D-R

64. D-tyr-ester-D-try-peptide-D-try-ester-L-R

65. D-tyr-ester-D-try-peptide-L-try-ester-D-R

66. D-tyr-ester-L-try-peptide-D-try-ester-D-R

67. L-tyr-ester-D-try-peptide-D-try-ester-D-R

68. D-tyr-ester-D-try-peptide-D-try-ester-D-R

69.L-R-ester-L-tyr-ester-L-try-peptide-L-try-ester-L-R

70.L-R-ester-D-tyr-ester-L-try-peptide-L-try-ester-L-R

71.L-R-ester-L-tyr-ester-D-try-peptide-L-try-ester-L-R

72.L-R-ester-L-tyr-ester-L-try-peptide-D-try-ester-L-R

73.L-R-ester-L-tyr-ester-L-try-peptide-L-try-ester-D-R

74.L-R-ester-D-tyr-ester-D-try-peptide-L-try-ester-L-R

75.L-R-ester-D-tyr-ester-L-try-peptide-D-try-ester-L-R

76.L-R-ester-D-tyr-ester-L-try-peptide-L-try-ester-D-R

77.L-R-ester-L-tyr-ester-D-try-peptide-D-try-ester-L-R

78.L-R-ester-L-tyr-ester-D-try-peptide-L-try-ester-D-R

79.L-R-ester-L-tyr-ester-L-try-peptide-D-try-ester-D-R

80.L-R-ester-D-tyr-ester-D-try-peptide-D-try-ester-L-R

81.L-R-ester-D-tyr-ester-D-try-peptide-L-try-ester-D-R

82.L-R-ester-D-tyr-ester-L-try-peptide-D-try-ester-D-R

83.L-R-ester-L-tyr-ester-D-try-peptide-D-try-ester-D-R

84.L-R-ester-D-tyr-ester-D-try-peptide-D-try-ester-D-R

85.D-R-ester-L-tyr-ester-L-try-peptide-L-try-ester-L-R

86.D-R-ester-D-tyr-ester-L-try-peptide-ltry-ester-L-R

87.D-R-ester-L-tyr-ester-D-try-peptide-L-try-ester-L-R

88.D-R-ester-L-tyr-ester-L-try-peptide-D-try-ester-L-R

89.D-R-ester-L-tyr-ester-L-try-peptide-L-try-ester-D-R

90.D-R-ester-D-tyr-ester-D-try-peptide-L-try-ester-L-R

91.D-R-ester-D-tyr-ester-L-try-peptide-D-try-ester-L-R

92.D-R-ester-D-tyr-ester-L-try-peptide-L-try-ester-D-R

93.D-R-ester-L-tyr-ester-D-try-peptide-D-try-ester-L-R

94.D-R-ester-L-tyr-ester-D-try-peptide-L-try-ester-D-R

95.D-R-ester-L-tyr-ester-L-try-peptide-D-try-ester-D-R

96.D-R-ester-D-tyr-ester-D-try-peptide-D-try-ester-L-R

97.D-R-ester-D-tyr-ester-D-try-peptide-L-try-ester-D-R

98.D-R-ester-D-tyr-ester-L-try-peptide-D-try-ester-D.R

99.D-R-ester-L-tyr-ester-D-try-peptide-D-try-ester-D-R

100.D-R-ester-D-tyr-ester-D-try-peptide-D-try-ester-D-R

The amino acid and hydroxy amino acid transporter compounds inaccordance with the invention can be administered to animals as a class,including man, and both large and small animals, by any conventionaladministration procedure applicable to lysine itself, including, forexample, oral administration, transdermal administration, transnasaladministration, inhalation, sublingual administration, rectaladministration and parenteral administration. The compounds can beadministered as such or with a nontoxic inert therapeutic carrier.

The amino acid and hydroxyamino acid transport compound can beadministered per se, or in association with a pharmaceuticallyacceptable diluent or carrier. The invention accordingly also provides apharmaceutical composition in dosage unit form comprising from 0.1 toabout 500 mg of amino acid and hydroxyamino acid transporter compound,per dosage unit, together with a pharmaceutically acceptable nontoxicinert carrier or diluent therefor.

The amino acid and hydroxyamino acid transporter compound can beadministered to the animal in an amount to provide therapeutic compoundin an amount sufficient to ameliorate the condition alleviated by thetherapeutic compound, and will depend upon the condition, the species ofanimal, and the weight of the animal. For example, in humanadministration, a dosage of amino acid and hydroxyamino acid transportercompound within the range from about 0.1 mg/kg to about 500 mg/kg perday. In the treatment of lower test animals, a similar dosage range istherapeutic. The upper limit of dosage is that imposed by toxic sideeffects, and can be determined by trial and error for the animal to betreated, including humans.

To facilitate administration, the amino acid and hydroxyamino acidtransporter compound can be provided in composition form, and preferablyin dosage unit form. While the compound can be administered per se, itis normally administered in conjunction with a pharmaceuticallyacceptable carrier thereof, which dilutes the compound and facilitateshandling. The term "pharmaceutically acceptable" means that the carrier(as well as the resulting composition) is sterile and nontoxic.

The carrier or diluent can be solid, semisolid, or liquid, and can serveas a vehicle, excipient, or medium for the lysine transporter compound.Exemplary diluents and carriers include lactose, dextrose, sucrose,sorbitol, mannitol, starches, gum acacia, calcium phosphate, mineraloil, cocoa butter, oil of theobroma, alginates, tragacanth, gelatin,syrup, methyl cellulose, polyoxyethylene sorbitan monolaurate, methyl-and propylhydroxybenzoate, talc or magnesium stearate.

For convenience in handling, the amino acid and hydroxyamino acidtransporter compound and carrier or diluent can be enclosed orencapsulated in a capsule, sachet, cachet, gelatin, paper or othercontainer, especially when intended for use in dosage unit. The dosageunits can for example take the form of tablets, capsules, suppositoriesor cachets.

The following Examples illustrate various forms of dosage units in whichthe amino acid and hydroxyamino acid transporter compound can beprepared:

Example 1

    ______________________________________                                        Tablet formulation      Mg/tablet                                             ______________________________________                                        Amino acid and hydroxyamino acid                                                                      15                                                    transporter compound                                                          Lactose                 86                                                    Corn starch (dried)     45.5                                                  Gelatin                 2.5                                                   Magnesium stearate      1.0                                                   ______________________________________                                    

The amino acid and hydroxyamino acid transporter compound is powered andpassed through a mesh sieve and well mixed with the lactose and 30 mg ofthe corn starch, both passed through a sieve.

The mixed powders are massed with a warm gelatin solution, prepared bystirring the gelatin in water and heating to form a 10% w/w solution.The mass is granulated by passing through a sieve, and the moistgranules dried at 40° C.

The dried granules are regranulated by passing through a sieve and thebalance of the starch and the magnesium stearate is added and thoroughlymixed.

The granules are compressed to produce tablets each weighing 150 mg.

Example 2

    ______________________________________                                        Tablet formulation    Mg/tablet                                               ______________________________________                                        Amino acid and hydroxyamino acid                                                                    100                                                     transporter compound                                                          Lactose               39                                                      Corn starch (dried)   80                                                      Gelatin               4.0                                                     Magnesium stearate    2.0                                                     ______________________________________                                    

The method of preparation is identical with that of Example 1 exceptthat 60 mg of starch is used in the granulation process and 20 mg duringtabletting.

Example 3

    ______________________________________                                        Capsule formulation   Mg/capsule                                              ______________________________________                                        Amino acid and hydroxyamino acid                                                                    250                                                     transporter compound                                                          Lactose               150                                                     ______________________________________                                    

The amino acid and hydroxyamino acid transporter compound and lactoseare passed through a sieve and the powders well mixed together beforefilling into hard gelatin capsules of suitable size, so that eachcapsule contains 400 mg of mixed powders.

Example 4

    ______________________________________                                        Suppositories        Mg/suppositories                                         ______________________________________                                        Amino acid and hydroxyamino acid                                                                    50                                                      transporter compound                                                          Oil of Theobroma     950                                                      ______________________________________                                    

The amino acid and hydroxyamino acid transporter compound is powderedand passed through a sieve and triturated with molten oil of theobromaat 45° C. to form a smooth suspension.

The mixture is well stirred and poured into molds, each of nominal 1 gcapacity, to produce suppositories.

Example 5

    ______________________________________                                        Cachets               Mg/cachet                                               ______________________________________                                        Amino acid and hydroxyamino acid                                                                    100                                                     transporter compound                                                          Lactose               400                                                     ______________________________________                                    

The amino acid and hydroxyamino acid transporter compound is passedthrough a mesh sieve, mixed with lactose previously sieved and fittedinto cachets of suitable size so that each contains 500 mg.

Example 6

    ______________________________________                                        Intramuscular injection                                                       (sterile suspension in aqueous vehicle)                                                              Mg                                                     ______________________________________                                        Amino acid and hydroxyamino acid                                                                     10                                                     transporter compound                                                          Sodium citrate         5.7                                                    Sodium carboxymethylcellulose                                                                        2.0                                                    Methyl para-hydroxybenzoate                                                                          1.5                                                    Propyl para-hydroxybenzoate                                                                          0.2                                                    Water for injection to 1.0 ml                                                 ______________________________________                                    

Example 7

    ______________________________________                                        Intraperitoneal intraveneous or                                               subcutaneous injection                                                        (sterile solution in aqueous carrier system)                                                           Mg                                                   ______________________________________                                        Amino acid and hydroxyamino acid                                                                       15                                                   transporter compound                                                          Sodium citrate           5.7                                                  Sodium carboxymethylcellulose                                                                          2.0                                                  (low viscosity grade)                                                         Methyl para-hydroxybenzoate                                                                            1.5                                                  Propyl para-hydroxybenzoate                                                                            0.2                                                  Water for injection to 1.0 ml                                                 ______________________________________                                    

Following administration of the amino acid and hydroxyamino acidtransporter compound, the metabolic system of animal, utilizing theactivity of esterases naturally present, will cleave the ester bondlinking the amino acid and hydroxyamino acid to the therapeuticcompound, and therefore make it available to exert its therapeuticeffect in the animal body. The therapeutic effectiveness of thecompounds following administration has been demonstrated by laboratorytests in small animals, as is evidenced by the following Example.

EXAMPLE 1

In this Example, the compound O-L-lysyl-L-tyrosine was employed as thetest compound.

Male CD-COBS rats (Charles River, Italy) weighing 200 g were used. Theanimals housed under standard conditions of light-dark cycle,temperature and humidity with free access to water and food.O-L-lysyl-L-tyrosine was dissolved in saline and injectedintraperitoneally (i.p.) and intravenously (i.v.) 2 ml/kg at theequimolar dose of 1.62 mmol/kg. L-tyrosine was suspended in 0.5%carboxymethylcellulose and injected only i.p. (because it is insolublein aqueous solvent) at the equimolar dose of 1.62 mmol/kg. Controlsreceived saline. Rates were killed by decapitation 30 and 120 min. aftertreatment. Plasma and brain were rapidly removed and kept at -20° C.until assay. L-tyrosine was assayed by electrochemical detection coupledwith HPLC according to Holman et al. (R. B. Holman and B. M. Snage,1983, J. Chromatogr. 262, 415-419) with minor modifications.

As shown in Table I, brain L-tyrosine concentrations were significantlyincreased by O-L-lysyl-L-tyrosine at both intervals considered. PlasmaL-tyrosine was significantly raised by only 30 min. after the i.v. dose.As shown in Table II, brain L-tyrosine concentrations were significantlyincreased after i.p. doses of L-tyrosine (p<0.05) andO-L-lysyl-L-tyrosine (p<0.01) at both intervals considered. PlasmaL-tyrosine was raised only after O-L-lysyl-L-tyrosine and only 30 min.after injection.

                  TABLE I                                                         ______________________________________                                        Plasma and brain levels of L-tyrosine after intravenous                       injection of O--L-lysil-L-tyrosine (1.62 mmol/kg) to rats                                   Tyrosine (μg/g ± S.E.)                                    Compound  Time (min)                                                                              Plasma        Brain                                       ______________________________________                                        Saline     0        11.21 ± 0.95                                                                             13.21 ± 0.63.sup.                        Lys-tyrosine                                                                             30       .sup. 46.80 ± 2.99.sup.1                                                                 49.02 ± 1.50.sup.1                       Lys-tyrosine                                                                            120       16.50 ± 4.01                                                                             31.57 ± 0.70.sup.1                       ______________________________________                                         .sup.1 p < 0.01 Dunnett's test                                                Data are the mean of five animals.                                       

                  TABLE II                                                        ______________________________________                                        Plasma and brain levels of L-tyrosine after intraperitoneal                   injection of equimolar doses of 1.62 mmol/kg of L-tyrosine                    and O--L-lysyl-L-tyrosine to rats                                                           L-tyrosine (μg/g ± S.E.)                                  Compound  Time (min)                                                                              Plasma        Brain                                       ______________________________________                                        Saline    --        11.27 ± 1.85                                                                           13.68 ± 0.83                               L-tyrosine                                                                               30       16.92 ± 2.06                                                                           20.22 ± 1.75.sup.1                         L-tyrosine                                                                              120       12.72 ± 2.23                                                                           21.77 ± 1.52.sup.1                         Lys-tyrosine                                                                             30       26.48 ± 1.64.sup.2+                                                                    54.04 ± 3.78.sup.2++                       Lys-tyrosine                                                                            120       14.59 ± 1.87                                                                           40.68 ± 2.21.sup.++                        ______________________________________                                         .sup.1 p < 0.05                                                                             Dunnett's test (vs controls)                                   .sup.2 p < 0.01                                                                .sup.+ p < 0.05                                                                             Tukey's test (vs L-tyrosine 30')                               .sup.++ p < 0.01                                                               p < 0.01 Tukey's test (vs L-tyrosine 120')                                   Data are the mean of four animals.                                        

The transporter compound of the invention that is composed of lysine andtryptophan linked by an ester linkage is of particular therapeuticinterest because it is a combination of two natural amino acids.Tryptophan is one of the twenty-two amino acids of animal protein, andnormally about 7 mg per kilogram of body weight is required in the dailydiet. It is a precursor to serine, one of the neurotransmitters, and isfound in man mainly in the intestines, platelets, and the brain. Serineis believed to have a role in certain mental disorders, includingendogenous and psychotic depression.

Tryptophan is hydrophobic because of the influence of its side chain.This hydrophobicity is overcome by the linkage with lysine, which ishydrophilic.

Lysine is another essential amino acid of interest, because its basicside chain NH₃ ⁺ often imparts a positive electrical charge to theprotein into which it has been incorporated, and also enhances itssolubility in water. Therefore, coupling with tryptophan has profoundeffects upon the bioavailability and therefore the clinical uses of thecombination in animal applications.

Tryptophan is used empirically in many parts of the world as a hypnotic,as a mild tranquilizer, and even as an antidepressant, either as asingle agent or in combination with other antidepressants andneuroleptics. Accordingly, the lysine-tryptophan compounds of theinvention, when used in combination with other drugs, can permit asubstantial lowering of the effect dose of the other drug, with fewerdose-related adverse reactions. This new safety margin with psychtropicsparing action can be particularly useful with neuroleptic agents.

The enhanced bioavailability of tryptophan in the lysinetryptophancompounds of the invention make it useful as a hypnotic, sedative,muscle relaxant, and antidepressant. There is no evidence to indicateinteraction between tryptophan and alcohol, indicating that the compoundcan be administered safely to people who consume alcohol. The compoundcan also be useful to dampen distortions of the biological clock, withfew, if any, side effects. The compound can also be administered toanimals for the treatment of neurotic disorders, distress, pain andanxiety, such as during travel.

In the claims, the term "amino acid" is used generically to encompassall amino acids having at least one amino and one carboxylic acid groupincluding hydroxyamino acids that in addition have at least one hydroxylgroup.

Having regard to the foregoing disclosure the following is claimed asthe inventive and patentable embodiments thereof:
 1. An amino acidtransporter compound in which amino acid as a carrier is linked via anhydrolyzably ester linkage to a therapeutic compound and having theformula:

    NH.sub.3.sup.+ --AA--COO--Z.sub.1                          I

in which AA is lysine Z₁ is a therapeutic compound linked via a hydroxylgroup thereof to a carboxylic acid group of the amino acid or a linkingcompound attached to --COO-- of the amino acid and to the therapeuticcompound.
 2. An amino acid transporter compound according to claim 1 inwhich Z₁ is tryptophane.
 3. An amino acid transporter compound accordingto claim 1 having the formula:

    NH.sub.3.sup.+ --AA--COO--Y.sub.1 --OOC--Z.sub.2           III

    NH.sub.3.sup.+ --AA--COO--Y.sub.2 --COO--Z.sub.1           IV

in which hydrolyzably Y₁ is a linking compound having two hydroxylgroups and Y₂ is a linking compound having one hydroxyl and onecarboxylic acid or acyl group.
 4. An amino acid transporter compoundaccording to claim 3 in which Y is a tyrisine-peptide-tyrosine group. 5.An amino acid transporter compound according to claim 4 in which Z₁ andZ₂ are tryptophane.
 6. A pharmaceutical composition in dosage unit formfor treating a condition for which the therapeutic compound iseffective, comprising an amount within the range from about 0.1 to about500 mg of an amino acid transporter compound according to claim 1 perdosage amount therapeutically effective to ameliorate the condition forwhich the therapeutic compound is effective, together with apharmaceutically acceptable nontoxic carrier or diluent therefor.
 7. Apharmaceutical composition is dosage unit form for treating a conditionfor which the therapeutic compound is effective, comprising an amountwithin the range from about 0.1 to about 500 mg of an amino acidtransporter compound according to claim 2 per dosage amounttherapeutically effective to ameliorate the condition for which thetherapeutic compound is effective, together with a pharmaceuticallyacceptable nontoxic carrier or diluent therefor.
 8. A pharmaceuticalcomposition in dosage unit form for treating a condition for which thetherapeutic compound is effective, comprising an amount within the rangefrom about 0.1 to about 500 mg of an amino acid transporter compoundaccording to claim 3 per dosage amount therapeutically effective toameliorate the condition for which the therapeutic compound iseffective, together with a pharmaceutically acceptable nontoxic carrieror diluent therefor.
 9. A process for treating a condition for which atherapeutic compound is effective, which comprises administering to awarm-blooded animal having the condition a therapeutic and relativelynontoxic amount of amino acid transporter compound according to claim 1,sufficient to ameliorate the condition.
 10. A process according to claim9 in which the amino acid transporter compound is administered in dosageunit form.
 11. A process according to claim 9 in which the amino acidtransporter compound is administered in an amount with the range fromabout 0.1 to about 500 mg per day.
 12. A process according to claim 9 inwhich the amino acid transporter compound is administered with an inertdiluent or carrier.
 13. A process according to claim 9 in which theamino acid transporter compound is administered orally.
 14. A processaccording to claim 9 in which the amino acid transporter compound isadministered parenterally.